ORALS

SESSION: OxidativeMonPM1-R1	Yoshikawa International Symposium (2nd Intl. Symp. on Oxidative Stress for Sustainable Development of Human Beings)
Mon. 28 Nov. 2022 / Room: Ballroom B
Session Chairs: Fuhua YANG; Yoshiaki Harakawa; Session Monitor: TBA

15:15: [OxidativeMonPM108] OS
Effect of antioxidant Twendee X on radiation-induced oxidative stress
Yoshiaki Harakawa¹ ; Fuhua Yang² ; Toshikazu Yoshikawa³ ; Haruhiko Inufusa⁴ ; ¹Division of Anti-oxidant Research, Gifu University, Gifu, Japan; ²Division of Anti-Oxidant Research, Gifu University, Gifu, Japan; ³Louis Pasteur Center, Kyoto, Japan; ⁴TIMA, Gifu, Japan;
Paper Id: 418 [Abstract]

It is known that when a living organism is exposed to radiation, water molecules in the body ionize, producing reactive oxygen species, which in turn cause inflammation.<br />Twendee X (TwX) is an antioxidant supplement consisting of eight ingredients (coenzyme Q10, vitamin C, vitamin B2, niacin, L-cystine, succinic acid, fumaric acid, and L-glutamine). We investigated whether TwX and other substances can inhibit oxidative stress generated by radiation.<br />C57BL/6J mice were orally administered the test substance TwX (15 mg/kg) for 3 days at 24-hour intervals, irradiated with 6 Gy of radiation on the fourth day, and blood samples were taken 72 hours later, and survival was observed as is. Vitamin C (15 and 150 mg/kg), Resveratrol, Astaxanthin, Protandim, and Pycnogenol were used as antioxidants for comparison to TwX. Plasma hydroperoxide level was measured as blood oxidative stress (d-ROMs test).<br />Increased d-ROMs test value by radiation was reduced by one third in the group treated with TwX (15 mg/kg) before irradiation for 3 days (t-test, p<0.05), and the oxidative stress level was lower in the group treated for 7 days than in normal mice. In addition, the administration of TWX suppressed radiation-induced deaths.<br />Vitamin C (150 mg/kg) and Astaxanthin treatment groups reduced d-ROMs test values, but the difference was not statistically significant. The low concentrations of vitamin C (15 mg/kg), Pycnogenol, and Protandim did not change the d-ROMs test values compared to the control group, and for the Resveratrol group, the d-ROMs test values increased compared to the control group. Furthermore, Twendee Mtcontrol (TwM), which added seven active ingredients to TwX, reduced radiation-related deaths with fewer doses than TwX.<br />TwX significantly reduced radiation-induced oxidative stress at a small dose of 15 mg/kg, while other single antioxidants with normal amount have no significant improvement in animal radiation model. High dose antioxidants may cause side effects. TwX, TwM are composed only of substances that exist in the human body, and a small dose of them showed high antioxidant activity. This may have reduced inflammation caused by oxidative stress, ultimately leading to an increase in survival rate. The antioxidant potential of Twendee series in this experiment suggests the possibility of future applications for various oxidative stress-related diseases.

References:
[1] Tadokoro K, et al. Clinical Benefits of Antioxidative Supplement Twendee X for Mild Cognitive Impairment: A Multicenter, Randomized, Double-Blind, and Placebo-Controlled Prospective Interventional Study. J Alzheimers Dis. 2019;71(3):1063-1069.\n[2] Yamamoto T, Kinoshita M, Shinomiya N, Hiroi S, Sugasawa H, Matsushita Y, Majima T, Saitoh D, Seki S. Pretreatment with ascorbic acid prevents lethal gastrointestinal syndrome in mice receiving a massive amount of radiation. J. Radiat. Res., 2010. 51, 145-156．\n[3] Yoshikawa T, et al. [All about Free Radicals – from Health to Environmental Pollution -] Kasseisanso , Free Radical no subete – Kennkou kara kannkyouosen made – (in Japanese), Maruzen Publishing, 2000, 78-79.\n[4] Seki Y. [Evaluation of total oxidative stress by d-ROMs testing] d-ROMs test niyoru sankasutoresu sougouhyouka (in Japanese). Seibutsu siryou bunseki (Journal of analytical bio-science), 2010. 32(4), 301-306.

SESSION: OxidativeTuePM1-R1	Yoshikawa International Symposium (2nd Intl. Symp. on Oxidative Stress for Sustainable Development of Human Beings)
Tue. 29 Nov. 2022 / Room: Ballroom B
Session Chairs: Shigeru Hirano; Session Monitor: TBA

14:25: [OxidativeTuePM106] OS
Role of Antioxidants in Oxidative Stress Induced by High Intake of Alcohol
Fuhua Yang¹ ; Yoshiaki Harakawa² ; Toshikazu Yoshikawa³ ; Haruhiko Inufusa⁴ ; ¹Division of Anti-Oxidant Research, Gifu University, Gifu, Japan; ²Division of Anti-oxidant Research, Gifu University, Gifu, Japan; ³Louis Pasteur Center, Kyoto, Japan; ⁴TIMA, Gifu, Japan;
Paper Id: 429 [Abstract]

Although there are genetic differences in the metabolism of alcohol, large amounts of alcohol intake can cause hangovers and, in severe cases, death due to acute alcohol poisoning. Once alcohol enters the body, it is metabolized mainly in the liver, where it is broken down into the highly hepatotoxic acetaldehyde. It is thought that excessive alcohol intake causes insufficient metabolism in the body, and the acetaldehyde induces oxidative stress, resulting in hangovers and organ damage. Therefore, we examined the effects of large amounts of alcohol on the body using Twendee S (TwS), an antioxidant formula consisting of coenzyme Q10, vitamin C, vitamin B2, niacin, L-cystine, succinic acid, fumaric acid, and L-glutamine. Mice were intraperitoneally administered ethanol, and blood acetaldehyde and oxidative stress were measured. The results showed that acetaldehyde in the blood increased at 2 hours after ethanol administration and was hardly detectable at 24 hours. However, oxidative stress remained high even after 72 hours of ethanol administration, and antioxidant capacity was low. In contrast, TwS-treated group suppressed blood acetaldehyde and the rising oxidative stress without reducing antioxidant capacity. In the mouse acute alcohol intoxication experiment, all mice in the Control group, to which nothing was administered, died after 12 hours. However, in the group that received 4 doses of TwS at 12-hour intervals the survival rate was 100%. These results suggest that heavy alcohol intake increases oxidative stress in the body, and that this oxidative stress may be a factor in acute poisoning deaths. In contrast, the antioxidant compound TwS suppressed acute poisoning deaths by speeding up the metabolism of alcohol and suppressing elevated oxidative stress. In conclusion, heavy and prolonged alcohol intake causes significant oxidative stress in the body, which may lead to aging and disease. Therefore, countermeasures against oxidative stress caused by alcohol intake are important, and TwS was shown to have the potential to reduce this risk.

References:
1). Markus Graf V. Matuschka-Greinffenclau, Hans Peter Jander. Alcohol metabolism moderating composition. WO 2005077464 A1. 2005-08-25.\n2). Markus Graf Matuschka Von Greiffenclau, Haruhiko Inufusa. Composition for accelerating alcohol metabolism and for reducing the risk of alcohol induced diseases. WO 2012095509 A8. 2012-10-04.\n3). Petrella C, Carito V Ceccanti M, et al. Oxidative stress inhibition by resveratrol in alcohol-dependent mice. Nutrition. 2020;79-80:110783.

SESSION: OxidativeTuePM1-R1	Yoshikawa International Symposium (2nd Intl. Symp. on Oxidative Stress for Sustainable Development of Human Beings)
Tue. 29 Nov. 2022 / Room: Ballroom B
Session Chairs: Shigeru Hirano; Session Monitor: TBA

14:50: [OxidativeTuePM107] OS
Potential for Antioxidant Therapy for Diabetes and Diabetic Complications
Yoshiaki Harakawa¹ ; Fuhua Yang² ; Toshikazu Yoshikawa³ ; Haruhiko Inufusa⁴ ; ¹Division of Anti-oxidant Research, Gifu University, Gifu, Japan; ²Division of Anti-Oxidant Research, Gifu University, Gifu, Japan; ³Louis Pasteur Center, Kyoto, Japan; ⁴TIMA, Gifu, Japan;
Paper Id: 426 [Abstract]

Diabetes is known to be a disease caused by oxidative stress, and also known to generates its own oxidative stress, resulting in numerous complications. Twendee X (TwX) is an antioxidant combination supplement with international and Japanese patents, and has been certified as effective in preventing dementia from a randomized, double-blind clinical trial led by the Japan Society for Dementia Prevention in 2019. In recent years, we have also successfully developed Twendee Mtcontrol (TwM), an antioxidant with seven more ingredients added to TwX to increase its antioxidant effect. The antioxidants TwX and TwM were tested in animal models to see if they can treat diabetes and diabetic complications. To examine the effect of antioxidants on diabetes, a diabetic disease model was developed. TwX or TwM was orally administered daily for 2 months. Afterwards, a glucose tolerance test was performed, and blood glucose levels were measured over time. In addition, insulin levels were measured under the same conditions. The results showed that the peak blood glucose level was significantly improved in the TwX group, and the peak level was also improved in the TwM group compared to the control group. Insulin levels also tended to be lower than controls in both groups. In addition, to verify the effectiveness of the supplements in preventing actual diabetic complications, we tested it on SDT fatty rats, which develop diabetic complications early in the course of the disease. These rats develop elevated blood pressure and cataracts at a much earlier stage due to the progression of diabetes mellitus. SDT fatty rats (5-week-old males) were divided into three groups (n=8): control (non-treated), TwX 40 mg/kg/Day, and TwM 40 mg/kg/Day. At 17 weeks of age, most rats in the non-treated group showed severe cataracts, whereas those in the TwM-treated group had relatively mild cataracts. Systolic blood pressure also tended to increase with age in the non-treated group, but the TwX and TwM groups significantly suppressed the age-related increase in blood pressure. These results suggest that TwX and TwM reduced blood glucose levels in an insulin-independent manner, thereby reducing diabetic complications such as hypertension and cataracts. TwX reduced blood glucose levels more, and TwM had a pronounced cataract-preventive effect. TwM may have a broader range of disease-preventive effects. however, further studies are required for the detailed mechanism of action. These results suggest the possibility of antioxidant therapy for diabetes mellitus and its complications.

References:
[1] Yamagishi S. [Diabetes and Oxidative Stress] Tounyoubyou to Sanka Sutoresu (in Japanese), Medical Review Co., Ltd., 2011, 3.\n[2] Masuyama T, Katsuda Y, and Shinohara M. A novel model of obesity-related diabetes: introgression of the Lepr^fa allele of the Zucker fatty rat into nonobese Spontaneously Diabetic Torii (SDT) rats. Exp. Anim. 54(1): 13-20, 2005.\n[3] Matsui K, Ota T, Matsushita M, et al. Diabetes-associated complications in Spontaneously Diabetic Torii Fatty Rats. Exp. Anim. 57(2): 111-121, 2008.

SESSION: OxidativeWedPM1-R1	Yoshikawa International Symposium (2nd Intl. Symp. on Oxidative Stress for Sustainable Development of Human Beings)
Wed. 30 Nov. 2022 / Room: Ballroom B
Session Chairs: Koji Fukui; Koji Abe; Session Monitor: TBA

14:00: [OxidativeWedPM105] OS Plenary
Twendee X improves cognitive function in vitamin E-deficient mice
Koji Fukui¹ ; Fuhua Yang² ; Yugo Kato¹ ; Yoshiaki Harakawa³ ; Toshikazu Yoshikawa⁴ ; Haruhiko Inufusa⁵ ; ¹Shibaura Institute of Technology, Saitama, Japan; ²Division of Anti-Oxidant Research, Gifu University, Gifu, Japan; ³Division of Anti-oxidant Research, Gifu University, Gifu, Japan; ⁴Louis Pasteur Medical Research Center, Kyoto, Japan; ⁵TIMA, Gifu, Japan;
Paper Id: 419 [Abstract]

Reactive oxygen species may attack several living organs and gradually accumulate oxidative products such as lipid hydroperoxides. Recently, several kinds of evidence have been demonstrated that accumulation of oxidative products induces onset and progression of several severe diseases such as inflammations, cardiovascular events and more [1]. One severe disease is dementia, and it also deeply relates to senescence. To prevent age-related neurodegenerative disorders, treatment with antioxidant is effective. Twendee X, a mixed antioxidant supplement contains 8 substances [2] and anyone can buy it. However, the detailed scientific evidence is not enough. In this study, we fed Twendee X to vitamin E-deficient mice and check cognitive function.<br />We created vitamin E-deficient mice and Twendee X treated for one month. Cognitive function was measured by the Morris Water maze task [3].<br />Vitamin E deficient mice were significantly decrease learning ability. Treatment with Twendee X was significantly improved it. Brain neurotrophic factors in Twendee X-treated mouse were significantly increase compared to the untreated groups.<br />Twendee X has a potential as a powerful antioxidant supplement through protection against neuronal dysfunction.

References:
[1] Yoshida, N, Kato, Y, Takatsu, H, et al., Biomedicines 10 (2022) 28.\n[2] Tadokoro, K, Morihara, R, Ohta, Y, et al., J Alzheimer’s Dis, 71 (2019) 1063-1069.\n[3] Fukui, K, Onodera, K, Shinkai, T, et al., Ann NY Acad Sci, 928 (2001) 169-176.

SESSION: OxidativeWedPM2-R1	Yoshikawa International Symposium (2nd Intl. Symp. on Oxidative Stress for Sustainable Development of Human Beings)
Wed. 30 Nov. 2022 / Room: Ballroom B
Session Chairs: Hironori Nakagami; Session Monitor: TBA

15:55: [OxidativeWedPM209] OS
Usefulness of Antioxidant Combination Products for Chronic Fatigue Syndrome
Fuhua Yang¹ ; Yoshiaki Harakawa² ; Toshikazu Yoshikawa³ ; Haruhiko Inufusa⁴ ; ¹Division of Anti-Oxidant Research, Gifu University, Gifu, Japan; ²Division of Anti-oxidant Research, Gifu University, Gifu, Japan; ³Louis Pasteur Center, Kyoto, Japan; ⁴TIMA, Gifu, Japan;
Paper Id: 427 [Abstract]

In Chronic Fatigue syndrome (CFS), a complex combination of factors such as infections and excessive stress in daily life causes abnormalities in the immune system, endocrine system, brain, and neurological functions. The pathogenesis of the syndrome is not yet clear, and no treatment has been established. Recently, it has been reported that CFS-like symptoms are also observed in COVID-19 sequelae and SARS-CoV-2 vaccine sequelae, and there is an urgent need to clarify the pathogenesis mechanism and establish prevention and treatment methods. Abnormal energy metabolism due to mitochondrial dysfunction and inflammation in the brain have been reported as one of the causes of CFS. Since oxidative stress is highly related to both of these factors, it was thought that suppressing oxidative stress may lead to improvement of various symptoms of CFS. Therefore, we analyzed the symptom changes before and after the use of an antioxidant combination supplement, Twendee Mtcontrol (TwM), taken by people who had been diagnosed with chronic fatigue syndrome. 2 months of TwM intake resulted not only in an anti-fatigue effect, but also in improvement of pains and QOL, compared to before intake. Although the effects of various antioxidants have been reported for CFS, most of them are mainly for fatigue, and there are no reports showing significant effects on various symptoms. This may be due to differences in the combination of antioxidants and their antioxidant capacity. The results of this study may be useful for future treatment and prevention of CFS.

References:
1). Feng T, Yamashita T, Abe K, et al. In Vitro Free Radical Scavenging Activities of Dietary Supplements by Electron Spin Resonance. Brain Supplements. 2020; 2: 1-12.\n2). You F, Tanaka S, Inufusa H, et al. Effects of Antioxidant composition Twendee X on side effects of SARS -COV-2 mRNA vaccine. Brain Supplement. 2022; 4: 1-6.\n3). You F, Tanaka S, Inufusa H, et al. Antioxidant composition Twendee X may improve long COVID symptoms. Brain Supplement. 2022; 4: 7-12.

SESSION: OxidativeWedPM2-R1	Yoshikawa International Symposium (2nd Intl. Symp. on Oxidative Stress for Sustainable Development of Human Beings)
Wed. 30 Nov. 2022 / Room: Ballroom B
Session Chairs: Hironori Nakagami; Session Monitor: TBA

16:20: [OxidativeWedPM210] OS
Antioxidants and Cancer Cells
Fuhua Yang¹ ; Yoshiaki Harakawa² ; Naomi Okada³ ; Toshikazu Yoshikawa⁴ ; Haruhiko Inufusa⁵ ; ¹Division of Anti-Oxidant Research, Gifu University, Gifu, Japan; ²Division of Anti-oxidant Research, Gifu University, Gifu, Japan; ³Gifu University, Gifu, Japan; ⁴Louis Pasteur Center, Kyoto, Japan; ⁵TIMA, Gifu, Japan;
Paper Id: 428 [Abstract]

Oxidative stress is involved in over 150 different diseases, and cancer is one of the most common. High oxidative stress can damage proteins and DNA that are essential to the body, leading to the formation of cancer over time. Oxidative stress in the body is also elevated in carcinoma carriers. When the body's oxidative stress is elevated, inflammatory substances increase, and the body's cancer immunity is reduced, which is thought to promote cancer cell growth and metastasis. Since antioxidants are thought to play an effective role in breaking this vicious cycle, we used the antioxidant compound Twendee X (TwX) to verify the effect of TwX on cancer cells through in vivo and in vitro experiments. Human colon cancer RPMI4788 cells are used as cancer cells. In in vitro experiments, the addition of 60 μg/ml of TwX to RPMI4788 cell culture medium slowed cell proliferation from 1 hour after addition, whereas the same concentration of the antioxidant Vitamin C did not inhibit cell proliferation. The inhibition of cell proliferation by the addition of TwX was not observed in normal cells. A mouse model of lung metastasis was created by transplanting RPMI cells (2 x 104 cells/ml) into nude mice via intravenous tail injection. After 6 weeks of treatment with TwX (40 mg/kg/day), blood samples were taken, and oxidative stress in the plasma, lung　metastasis, and NK activity was measured. The results showed that the oxidative stress level of the tumor-bearing mice was twice as high as that of normal mice, but TwX suppressed the increase from the normal level to less than half. Lung weights were also significantly lighter than controls. NK activity was significantly lower NK activity than normal mice, while the TWX group had twice higher NK activity as the carriers and 1.5 times higher than the normal mice. These results suggest that TwX may suppress cancer metastasis by suppressing elevated oxidative stress, thereby increasing NK activity, which is cancer immunity. However, similar results may not be obtained in the case of single-component antioxidants. We believe that the results of this study will help antioxidants in cancer treatment and prevention.

References:
1. Composition for protection against cell-damaging effects. WO/2013/072441 23.05.2013 (patent)\n2. Novel compositions and uses thereof WO 2008071790 A3\n3. Suplatast tosilate reduces radiation-induced lung injury in mice through suppression of oxidative stress. Yusuke Izumi, Taku Nakashima, Noboru Hattori, Free Radic Biol Med.2019;136:52-59